A long Stokes shift red fluorescent Ca2+ indicator protein for two-photon and ratiometric imaging.

ABSTRACT: The introduction of calcium ion (Ca(2+)) indicators based on red fluorescent proteins (RFPs) has created new opportunities for multicolour visualization of intracellular Ca(2+) dynamics. However, one drawback of these indicators is that they have optimal two-photon excitation outside the near-infrared window (650-1,000?nm) where tissue is most transparent to light. To address this shortcoming, we developed a long Stokes shift RFP-based Ca(2+) indicator, REX-GECO1, with optimal two-photon excitation at <1,000?nm. REX-GECO1 fluoresces at 585?nm when excited at 480?nm or 910?nm by a one- or two-photon process, respectively. We demonstrate that REX-GECO1 can be used as either a ratiometric or intensiometric Ca(2+) indicator in organotypic hippocampal slice cultures (one- and two-photon) and the visual system of albino tadpoles (two-photon). Furthermore, we demonstrate single excitation wavelength two-colour Ca(2+) and glutamate imaging in organotypic cultures.

SUBMITTER: Wu J 

PROVIDER: S-EPMC4920544 | biostudies-literature | 2014 Oct

REPOSITORIES: biostudies-literature